It has been more than a century since a virus was used to treat neoplastic disease. Doctors noticed that occasionally a patient was cured after a virus infection. This puzzle was not resolved until the beginning of 1920s and since then attracted attentions from scientists and clinical doctors. That was the beginning of the virotherapy (using viruses to treat cancer). By 1950s, more than 50 viruses had been tested for anti-tumor activity in animals or patients (Mullen and Tanabe (2002) The Oncologist 7:106-119; McCormick F (2001) Nature Review Cancer 1:130-141). In 1956, Dr. Smith and his team treated more than 30 cervical cancer patients with lysate of HeLa cells or KB cells that were infected with 10 serotypes of wild-type adenoviruses through intratumor injection, intrahepatic arterial injection or intravenous injection. No severe side effects were observed except the flu symposium in most patients; on the other hand, tumors of several patients went shrinkage and were necrotic. Unfortunately, due to the limitation of virus production band purification, this work did not proceed further. Dr. Smith's work had encouraged many scientists to explore the possibility of using viruses to treat cancers (Chiocca E A (2002) Nature Review Cancer 2: 938-950).
Along with the progress made in molecular biology and genetic engineering, in particular, with better understandings into viruses in relation to human being, scientists have been enabled to genetically manipulate viral genome since late last century, which includes the success of creating virus mutants that can specifically replicate in tumor cells (McCormick F (2001) Nature Review Cancer 1:130-141). For example, glioma-specific herpes virus mutant G207 (Martuza et al., (1991) Science 252:854-856), adenovirus Addl1520 (Onyx-015), an adenovirus mutant preferentially replicating in p53 defective tumor cells (Bischoff et al., (1996) Science 274:373-376) and CV706, a prostate cancer cell specific adenovirus mutant (Rodriguez et al. (1997) Cancer Research 57:2559-2563). This work has laid a foundation for the formation of new field: Cancer Virotherapy. At present, there are over 20 virus variants in clinical trials (Mullen and Tanabe (2002) The Oncologist 7:106-119).
There are several approaches for making tumor cell-specific oncolytic viruses. One approach to make a virus variant that selectively replicates in tumor cells is to use tumor cell-specific regulatory elements such as promoters and enhancers to control expression of essential viral genes, for example, the E1A, E1B, E2 and E4 genes of adenovirus (DeWeese et al. (2001) Cancer Research 61:7464-7472). By this approach, essential viral genes and ultimately virus replication would be under the control of tumor cell-specific regulatory elements, such regulatory elements include prostate-specific antigen (PSA) promoter and enhancers, alpha-feto protein (AFP) promoter and enhancers, human E2F-1 promoter etc. (McCormick F (2001) Nature Review Cancer 1:130-14).
Telomerase is an important enzyme for controlling length of cells' chromosome end, capable of regulating the length of the chromosome end during the process of cell fission. Telomerase is consisted mainly of the three parts, wherein RNA and telomerase reverse transcriptase gene (hTERT), which is of catalytic activity, control the activity of telomerase, while hTERT promoter determines the expression and activity of the telomerase.
Further researches have indicated that telomerase has no or very low activity in adult's normal cells. (Kim N W et al. Science. Dec. 23, 1994; 266(5193):2011-5; Shay J W et al. European Journal of Cancer 1997, 33;271-282), while it has a high delivery level in more than 90% tumor cells (Hahn and Weinberg (2002) Nature Review Cancer 2:331-341; Shay and Wright (1996) Current Opinion Oncology 8:66-71). Based on these features of hTERT, scientists have successfully generated several vectors in which the hTERT promoter was incorporated into the viral genome for gene delivery, including the works recently published describing the use of the hTERT promoter for conditionally replicating oncolytic adenoviruses (Lanson et al. (2003) Cancer Research 63:7936-7941; Kawashima et al., (2004) Clinical Cancer Research 10:285-292; Kim et al. (2003) Oncogene 22:370-380; Irving et al. (2004) Cancer Gene Therapy 11:174-185).
There are a few endogenous regulatory elements in adenoviral genome, which regulates expression of viral genes, for example, the promoter and enhancer of the E1A gene, among these endogenous regulatory elements, the sequence of the E1A enhancer overlaps with the viral packaging signal. In order to minimize the impact of endogenous regulatory elements on the regulation of a heterologous promoter, scientists had relocated the packaging signal to the right arm from the native site at the left end (Bristol et al. (2003) Molecular Therapy 7(6):755-764; Jakubczak et al. (2003) Cancer Research 63:1490-1499). Unfortunately, this kind of relocation of viral packaging signal from the native site to the right end has destabilized viral genome, resulting in the generation of many viral mutants (WO 02/067861; ASGT 2003 Annual Meeting, Molecular Therapy). Therefore, the above said relocation of viral packaging signal is not a viable approach to minimize the impact of endogenous regulatory elements. How to minimize the impact of the endogenous viral regulators on the heterologous elements and to prevent undesired generation of many viral mutants, while keep the viral genome stable?
With respect to the expression level of telomerase in humans, during the embryonic development and differentiation, scientists have also come to recognize that there is a certain level of telomerase activity in some cells including cervical member cells, progenitors, and stem cells (Wright et al. (1996) Development Genetics 18:173-179; Sharma et al. (1995) Proc. Natl. Acad. Sci. USA 92:12343-12346; Kolquist et al. (1996) British Journal of Cancer 80:1156-1161; Tahara et al. (1999) Oncogene 18:1561-1567; Tanaka et al. (1998) American Journal of Pathology 153:1985-1991). Low level of telomerase expression activity has cautioned scientists when they used the hTERT promoter to deliver a therapeutic gene in gene therapy vector, or to generate tumor cell-specific oncolytic adenoviruses by controlling essential viral genes (Hahn WC (2004) Clinical Cancer Research 10:1203-1205). Due to low level of telomerase expression in non-targeting cells, such as the progenitors, the hTERT promoter controlling oncolytic adenoviruses may replicate massively in progenitor cells, resulting in severe side effects (Huang et al. (2004) Clinical Cancer Research 10:1439-1445; Hahn WC (2004) Clinical Cancer Research 10:1203-1205; Masutomi et al. (2003) Cell 114:241-253). Therefore, increasing tumor cell specificity of the hTERT promoter is one of the key projects in generating oncolytic adenovirus using the hTERT promoter.